The research combines computer simulation (energy optimization/expansion techniques, Monte-Carlo calculations, and normal-mode analysis) with a new naturally discrete model of DMAto examine the configurations and properties of supercoiled DMA,a topologically constrained form of the double helix subject to higher-order folding and compensatory strand twisting and the way that DMAexists in living cells. Base sequence-dependentfeatures of the long, threadlike polymer are incorporated in the theory and treated by numerical simulations. By combining analytical studies with computer simulations, we obtain complementary information and have a series of built-in checks and balances for assessing the significance of our findings. The computational results stimulate new theoretical developments, which in turn can be used to assess the validity of the calculations. The effects of the sugar-phosphate backbone and local chemical environment are treated implicitly with knowledge-based potentials extracted from high-resolution structures of double-helical DMAand, in representative cases, with explicit treatment of long-range forces. Our immediate goals are to study the sequence-dependent biophysical properties of DMA minicircles and loops and to establish the physico-chemical basis of in-vivo looping. The proposed studies aim to clarify the role of primary chemical features (base sequence), ligand binding (proteins), and levels of supercoiling (imposed constraints on base-pair positioning) on the overall folding of the double helix. We will extend our studies of DMA loops tethered to the Lac represser protein with the aim of gathering new insights into the role of the molecular assembly in the regulation of transcription and its potential applicability as a model of eukaryotic insulators. A second goal is to gain insight into the effect of DMAsequence and supercoiling on protein binding and consequent cellular function. Other issues to be addressed include: (1) the role of local sequence-dependent structure and ligand binding on large-scale configurational transitions of spatially constrained duplexes;(2) the competing effects of multiple proteins on the configurational properties of supercoiled molecules;and (3) the interplay of local and global structure in supercoiling dynamics.